MOLECULAR CLONING AND CHARACTERIZATION OF A SID‐1‐LIKE GENE IN Plutella xylostella

Abstract

RNA interference (RNAi) signal can spread from the point where the double-stranded RNA (dsRNA) was initially applied to other cells or tissues. SID-related genes in Caenorhabditis elegans help in the spreading of this signal. However, the mechanisms of systemic RNAi are still not unveiled in insects. In this study, we cloned a full-length cDNA of sid-1-like gene, Pxylsid-1, from Plutella xylostella that contains 1,047 bp opening reading frame encoding a putative protein of 348 amino acids. This transcript is very much similar to the sil-1 in Bombyx mori (68.8%). The higher expression levels of Pxylsid-1 were found at the adult and fourth-instar stages compared to the second-instar stage with 21.48- and 10.36-fold increase, respectively. Its expression levels in different tissues were confirmed with the highest expression in the hemolymph, which showed 21.09-fold increase than the midgut; however it was lower in other tissues. The result of RNAi by feeding bacterially expressed dsRNA targeting Pxylace-1, which showed that the mRNA level of Pxylace-1 decreased by 34.52 and 64.04% after 36- and 72-h treatment, respectively. However, the mRNA level of Pxylsid-1 was not significantly induced when the Pxylace-1 was downregulated. Furthermore, we found that downregulation of Pxylsid-1 did not affect the RNAi effect of Pxylace-1. Hence, the Pxylsid-1 may not be involved in absorption of dsRNA from the midgut fluid. A further study is needed to uncover the function of Pxylsid-1.

Movement of regulatory RNA between animal cells

“…Consistent with this possibility, knockdown of the SID-1-like protein in the migratory locust Locusta migratoria do not result in a defect in systemic RNAi (Luo et al, 2012). Nor do SID-1-like proteins in the diamondback moth Plutella xylostella (Wang et al, 2014) or in the red flour beetle Tribolium castaneum (Miller et al, 2012) appear to be required for RNAi. In planaria, silencing in response to ingested dsRNA can occur in most tissues (Rouhana et al, 2013) and weak homologs of SID-1 are present (Zayas et al, 2005) but the roles of these homologs in the uptake of dsRNA are yet to be evaluated.…”

Silencing of ecdysone receptor, insect intestinal mucin and sericotropin genes by bacterially produced double‐stranded RNA affects larval growth and development in Plutella xylostella and Helicoverpa armigera

“…Successful use of this class of RNA was henceforth reported in various insects, including Spodoptera exigua (Tian et al ., ), H. armigera (Zhu et al ., ; Zhang et al ., ), Bactrocera dorsalis (Li et al ., ) and Leptinotarsa decemlineata (Zhu et al ., ), amongst others. Recently, successful use of the IPTG inducible L4440 vector was reported in P. xylostella for functional characterization of the acetylcholinesterase and sid‐1 like genes (Wang et al ., ). In the present study, two related lepidopterans were chosen for the analysis, owing to a good degree of conservation of genes between the two species.…”

“…Wang et al . () for the first time reported the leaf coating method for carrying out feeding assays in P. xylostella targeting the insect acetylcholinesterase gene.…”

The mysteries of insect RNAi: A focus on dsRNA uptake and transport

“…The identification of SID-1-like (SIL) proteins that include SILA, SILB, and SILC in multiple insect species has spurred several studies aiming to characterize their function in the uptake of dsRNA. The findings of these studies have ranged from supporting evidence for SIL protein function in RNAi response in WCR and CPB (Miyata et al, 2014;Cappelle et al, 2016), to no evidence for their involvement in desert locust, Schistocerca gregaria and diamondback moth, Plutella xylostella (Luo et al, 2012;Wang et al, 2014). The interpretations of the work that provides evidence for SILA and SILC involvement in RNAi include hypotheses for partial participation and redundant pathways for dsRNA uptake (Miyata et al, 2014;Cappelle et al, 2016).…”